Synthesis of Oligonucleotides Containing the (4R) and (4S) Diastereoisomers of 4,8-Dihydro-4-hydroxy-8-oxo-2′-deoxyguanosine

Abstract

The insertion of the (4R) and (4S) diastereoisomers of 4,8‐dihydro‐4‐hydroxy‐8‐oxo‐2′‐deoxyguanosine, the two main singlet oxygen oxidation products of 2′‐deoxyguanosine, (4R)‐4‐OH‐8‐oxodGuo (3a) and (4S)‐4‐OH‐8‐oxodGuo (3b), into oligonucleotides has been achieved by means of phosphoramidite chemistry using the solid‐phase synthesis approach. The synthesis of the phosphoramidite synthons 8a and 8b required 6 steps from 2′‐deoxyguanosine and involved the simultaneous protection of the additional tertiary hydroxy (4‐OH) and N2‐amino groups of the modified base by acetylation. The modified phosphoramidites 8a, b were efficiently incorporated into several oligonucleotides (3‐mer, 9‐mer, 14‐mer, and 22‐mer). The presence and the integrity of the 3a and 3b in the synthetic oligomers was confirmed by electrospray ionization mass spectrometry, together with HPLC and MALDI‐TOF mass‐spectrometric analyses of enzymatic digestions. The use of various enzymes, including endonucleases (nuclease P1) and exonucleases (snake venom phosphodiesterase and calf spleen phosphodiesterase), clearly shows that the enzymatic hydrolysis of phosphodiester bonds between 3a,b and normal 2′‐deoxyribonucleosides is inhibited.

Chemical Synthesis and Biochemical Properties of Oligonucleotides that Contain the (5′S,5S,6S)-5′,6-Cyclo-5-hydroxy-5,6-dihydro-2′-deoxyuridine DNA Lesion

“…Nuclease P 1 was found to be unable to cleave the phosphodiester bonds between normal and altered 2'-deoxyribonucleosides. [18,19] Enzymatic digestion of oligonucleotides containing (5'S,5S,6S)-cyclo-5-OH-dHdU (1) by bovine intestinal mucosa phosphodiesterase (3'-exo) and calf spleen phosphodiesterase (5'-exo): Additional enzymatic digestion experiments were performed on the modified 3-mer 11 by using the two exonucleases bovine intestinal mucosa phosphodiesterase (3'- exo) and calf spleen phosphodiesterase (5'-exo). It was shown that both enzymes failed to cleave the phosphodiester bond on either side of the (5'S,5S,6S)-cyclo-5-OHdHdU (1) residue, as already observed for (5'S,6S)-cyclodHdU and (5'S,6S)-cyclodHT.…”

Synthesis and Characterization of Oligonucleotides Containing 5‘,8-Cyclopurine 2‘-Deoxyribonucleosides: (5‘R)-5‘,8-Cyclo-2‘-deoxyadenosine, (5‘S)-5‘,8-Cyclo-2‘-deoxyguanosine, and (5‘R)-5‘,8-Cyclo-2‘-deoxyguanosine

“…However, the hydrolysis of the phosphodiester bonds between 2-4 and normal 2′-deoxyribonucleosides is much slower than that of the corresponding nonmodified ODNs, requiring 26 h of incubation with nuclease P1. Such results suggest a partial inhibition of the endonuclease activity by the lesion in agreement with previously reported data involving short ODNs containing other free radical-and singlet oxygen-induced base modifications (31)(32)(33). To obtain more information about the effect of specific tandem DNA lesions 2-4 on the behavior of nucleases, we have also studied the stability of the modified ODNs toward exonucleases.…”

Quantitation of Four Guanine Oxidation Products from Reaction of DNA with Varying Doses of Peroxynitrite

“…Cadet, et al, reported that enzymatic hydrolysis of phosphodiester bonds between 4,8-dihydro-4-hydroxy-8-oxo-dG (later to be corrected as spiroiminodihydantoin (13) and normal 2'-deoxyribonucleosides in oligonucleotides was inhibited with both endonucleases and exonucleases (33). In order to evaluate the effects of various enzymes on the formation of 8-oxodG and spiroiminodihydantoin, untreated calf thymus DNA was digested with 1.)…”